DISTRIBUTION OF CALRETININ IMMUNOREACTIVITY IN THE MOUSE DENTATE GYRUS .2. MOSSY CELLS, WITH SPECIAL REFERENCE TO THEIR DORSOVENTRAL DIFFERENCE IN CALRETININ IMMUNOREACTIVITY

In our previous study we revealed the presence oi clustered large calr etinin-immunoreactive multipolar cells in the ventral hilus of the mou se dentate gyrus and indicated that they might be messy cells, the pri ncipal neurons in the dentate hilus. In the present study we confirmed this identification with several methods and analysed further in deta il. In Golgi-impregnated samples messy cells were easily identified by their locations and characteristic thorny excrescences on their proxi mal dendrites. Golgi-impregnated messy cells were observed not only in the ventral hilus but also in the dorsal hilus, where no calretinin-i mmunoreactive large multipolar cells were encountered. Interestingly, messy cells exhibited dorsoventral differences in the size and complex ity of thorny excrescences; messy cells at the dorsal and middle level s had larger and more complex thorny excrescences, which covered dendr itic shafts for a longer distance, while ventral mossy cells had small er, simpler and shorter thorny excrescences. Confocal laser scanning l ight microscopic observations at a high magnification showed that the vast majority of calretinin-immunoreactive large neurons in the ventra l hilus displayed the thorny excrescences characteristic to messy cell s. Messy cells identified with the intracellular injection of Lucifer Yellow were calretinin-immunoreactive. Electron microscopic observatio ns clearly revealed that calretinin-immunoreactive elements showed str uctural features of messy cells such as thorny excrescences receiving typical synapses from messy fibre terminals. At the supragranular zone , a well-known target zone of messy cell axons, a dense calretinin-imm unoreactive band was seen, where numerous calretinin-immunoreactive pu nctae and fibres were packed. Electron microscopic observations reveal ed that these calretinin-immunoreactive axon terminals in the supragra nular zone made asymmetrical synapses on presumed granule cell dendrit ic spines. Tracer injection studies and lesion experiments indicated t hat the supragranular calretinin-immunoreactive axon terminals mainly originated from the large calretinin-immunoreactive multipolar cells i n the ipsilateral ventral hilus. Fluorescent double immunostaining for calretinin and glutamate receptor 2/3 (GluR2/3) revealed that all lar ge calretinin-immunoreactive hilar cells in the ventral level were Glu R2/3-immunoreactive and almost all intensely GluR2/3-immunoreactive hi lar cells in the ventral level were calretinin-immunoreactive. In addi tion intensely GluR2/3-immunoreactive but calretinin-negative large ce lls were encountered in the dentate hilus at the dorsal level. On the basis of these observations, we concluded that large calretinin-immuno reactive cells in the ventral hilus of. the mouse dentate gyrus were r eally messy cells and that messy cells at the dorsal level were calret inin negative. The present study revealed that mouse messy cells show the dorsoventral difference in the calretinin immunoreactivity and thu s they are chemically heterogeneous. (C) 1997 IBRO. Published by Elsev ier Science Ltd.